The 39 kD receptor-associated protein (RAP) is an ER-resident protein that was originally identified because it co-purified with the low-density lipoprotein related protein (LRP1) (Strickland et al., 1991 J. Biol. Chem. 266:13364-13369). Subsequent functional studies established that RAP plays a role in ensuring efficient delivery of LRP1 and other transmembrane proteins of the low-density lipoprotein receptor family of proteins (LDLRFP) to the cell surface (Lee et al., 2006 Moll Cell 22: 423-430). In addition, through binding to proteins of the LDLRFP, RAP can deliver therapeutic proteins and polypeptides into cells and across the blood brain barrier (Pan et al., 2004 J Cell Science 117: 5071-5078). Furthermore, RAP can attenuate the function of proteins of the LDLRFP, which can result in the suppression of cell activity and an increase of the in vivo half-life of LDLRFP ligands.
RAP consists of three domains of approximately 100 residues each. The affinity of the third domain of RAP (RAP-D3) for LRP1 is at least an order of magnitude greater than that of the other RAP domains. At neutral pH, wild-type (WT) RAP-D3 is highly helical, but it undergoes a cooperative thermal unfolding transition at 42° C., and a pH-induced unfolding transition at a pH of 6.3, and loses its ability to bind to members of the LDLRFP, including LRP1.
There remains a need for stabilized variants of RAP for improved transport of therapeutic compounds and therapeutic polypeptides into cells and across the blood brain barrier. In addition, there remains a need for stabilized variants of RAP to suppress cell activity and to increase the in vivo half-life of LDLRFP ligands.